1,25(OH)(2)D(3) stimulates Mg2+ uptake into MDCT cells: modulation by extracellular Ca2+ and Mg2+

The distal convoluted tubule plays a significant role in renal magnesium conservation. Although the cells of the distal convoluted tubule possess the vitamin D receptor, little is known about the effects of 1alpha,25-dihydroxyvitamin D [1,25(OH)(2)D(3)] on magnesium transport. In this study, we examined the effect of 1,25(OH)(2)D(3) on distal cellular magnesium uptake and the modulation of this response by extracellular Ca2+ and Mg2+ in an immortalized mouse distal convoluted tubule (MDCT) cell line. MDCT cells possess the divalent cation-sensing receptor (CaSR) that responds to elevation of extracellular Ca2+ and Mg2+ concentrations to diminish peptide hormone-stimulated Mg2+ uptake. Mg2+ uptake rates were determined by microfluorescence in Mg2+ -depleted MDCT cells. Treatment of MDCT cells with 1,25(OH)(2)D(3) for 16-24 h stimulated basal Mg2+ uptake in a concentration-dependent manner from basal levels of 164 +/- 5 to 210 +/- 11 nM/s, representing a 28 +/- 3% change. Pretreatment with actinomycin D or cycloheximide abolished 1,25(OH)(2)D(3)-stimulated(.)Mg2+ uptake (154 +/- 18 nM/s), suggesting that 1,25(OH)(2)D(3) stimulates Mg2+ uptake through gene activation and protein synthesis. Elevation of extracellular Ca2+ inhibited 1,25(OH)(2)D(3)-stimulated Mg2+ uptake (143 +/- 5 nM/s). Preincubation of the cells with an antibody to the CaSR prevented the inhibition by elevated extracellular Ca2+ of 1,25(OH)(2)D(3)-stimulated Mg2+ uptake (202 +/- 8 nM/s). Treatment with an antisense CaSR mRNA oligodeoxynucleotide also abolished the effects of extracellular Ca2+ on 1,25(OH)(2)D(3)-responsive Mg2+ entry. This showed that elevated extracellular calcium modulates 1,25(OH)(2)D-mediated responses through the CaSR. In summary, 1,25(OH)(2)D(3) stimulated Mg2+ uptake in MDCT cells, and this is dependent on de novo protein synthesis. Elevation of extracellular Ca2+, acting via the CaSR, inhibited 1,25(OH)(2)D(3)-stimulated Mg2+ entry. These data indicate that 1,25(OH)(2)D(3) has important effects on the control of magnesium entry in MDCT cells and these responses can be modulated by extracellular divalent cations.